Project Summary/Abstract Early and accurate diagnosis of disease is of paramount importance for positive prognosis. Point-of-care testing, defined as testing at or near the site of patient care, is an attractive alternative to laboratory-based testing. By providing results immediately, point-of-care diagnostic devices increase the expediency with which patients can proceed along the proper course of treatment. The goal of this project is to develop a low-cost point-of-care device to be used for the diagnosis of infectious diseases. The development of such a device that is also both sensitive and selective is desirable, yet challenging. We will employ paper-based diagnostics for the direct detection of pathogen-derived enzymes. Specifically, we aim to use fluorogenic peptidic substrates of bacterial/viral proteases (enzymes that catalyze the hydrolysis of peptide bonds) to detect infection by these agents. En route to this device, we envision a multistep process in which issues of sensitivity, selectivity, and reliability will be encountered and addressed systematically. This method of diagnosis should be applicable to any disease in which an enzyme is either misregulated (neoplastic and autoimmune diseases) or merely present (infection). We will focus our initial efforts on the creation of a diagnostic device for the detection of human immunodeficiency virus (HIV) infection. Twenty-five years ago it was discovered that acquired immunodeficiency syndrome (AIDS) is caused by HIV. Despite that knowledge, no cure has yet been developed. As a result, considerable time and effort has been spent developing tools to diagnosis HIV infection. Several immunoassay-based tests have been approved by the FDA, each with their own strengths and weaknesses. In low-resource settings, where HIV is highly prevalent, most of these tests remain restrictively expensive. Thus, the development of a rapid, low-cost test for HIV infection remains an important goal.